Publikationen - Molekulare Signalverarbeitung

Ribonuclease LE (RNaseLE) from tomato (Lycopersicon esculentum Mill. cv. Lukullus) belongs to the widespread RNase T2 family of ribonucleases. With the exception of S-RNases of the solanaceous self-incompatibility system the functions of other members of the RNase T2 family are only barely understood. Using a 2.6-kbp putative promoter sequence of RNaseLE in front of the uidA reporter gene, expression of β-glucuronidase in developing phloem tissue and, especially, in the meristematic and elongation zones at root tips was detected. The tissue-specific expression accords with the range of cis-acting elements detected in the RNaseLE promoter. RNaseLE mRNA was localized in developing phloem cells but not in mature phloem tissue, suggesting association of RNaseLE expression with phloem development. Histochemical staining of β-glucuronidase activity as well as detailed inspection of RNaseLE at mRNA, protein and enzyme activity levels revealed that the wound-induced expression of RNaseLE was also restricted to vascular tissue. RNaseLE transcript accumulation detected by in situ hybridization occurred preferentially in phloem and cambial cells of stem sections upon wounding. The data provide evidence for a role of RNaseLE in a tissue-specific wound response and in wound healing of tomato.

In this paper we report the in-planta activity of the ribosome-inactivating protein JIP60, a 60-kDa jasmonate-induced protein from barley (Hordeum vulgare L.), in transgenic tobacco (Nicotiana tabacum L.) plants. All plants expressing the complete JIP60 cDNA under the control of the cauliflower mosaic virus (CaMV) 35S promoter exhibited conspicuous and similar phenotypic alterations, such as slower growth, shorter internodes, lanceolate leaves, reduced root development, and premature senescence of leaves. Microscopic inspection of developing leaves showed a loss of residual meristems and higher degree of vacuolation of mesophyll cells as compared to the wild type. When probed with an antiserum which was immunoreactive against both the N- and the C-terminal half of JIP60, a polypeptide with a molecular mass of about 30 kDa, most probably a processed JIP60 product, could be detected. Phenotypic alterations could be correlated with the differences in the detectable amount of the JIP60 mRNA and processed JIP60 protein. The protein biosynthesis of the transformants was characterized by an increased polysome/monosome ratio but a decreased in-vivo translation activity. These findings suggest that JIP60 perturbs the translation machinery in planta. An immunohistological analysis using the JIP60 antiserum indicated that the immunoreactive polypeptide(s) are located mainly in the nucleus of transgenic tobacco leaf cells and to a minor extent in the cytoplasm.

The accumulation of abundant proteins and their respective transcripts, induced by 10−4 M cisabscisic acid or 10−5 M jasmonic acid methyl ester, was studied in barley (Hordeum vulgare L.) leaf segments and compared to that resulting from osmotic stress caused by floating the segments on solutions of sorbitol, glucose, polyethyleneglycol (PEG)-6000 or NaCl. Osmotic stress or treatment with abscisic acid led to the synthesis of novel proteins which were identical to jasmonateinduced proteins (JIPs) with respect to immunological properties and molecular masses. The most prominent polypeptides were characterized by molecular masses of 66, 37 and 23 kDa and were newly synthesized. Whereas sorbitol, mannitol, sucrose, glucose and PEG provoked the synthesis of JIPs, 2deoxyglucose and NaCl did not. We provide evidence that the synthesis of JIPs induced by osmotic stress is directly correlated with a preceding rise in endogenous jasmonates. These jasmonates, quantified by an enzyme immunoassay specific for (−)jasmonic acid and its aminoacid conjugates, increased remarkably in leaf segments treated with sorbitol, glucose or other sugars. In contrast, no increase in jasmonates could be observed in tissues exposed to salts (NaCl). The results strengthen the hypothesis that the accumulation of jasmonates, probably by de-novo synthesis, is an intermediate and essential step in a signalling pathway between (osmotic) stress and activation of genes coding for polypeptides of high abundance.